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MB Sample ID: SA127154
Local Sample ID: | 449_C |
Subject ID: | SU001588 |
Subject Type: | Other |
Subject Species: | Marine Plankton |
Gender: | Not applicable |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU001588 |
Subject Type: | Other |
Subject Species: | Marine Plankton |
Gender: | Not applicable |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
449_C | SA127154 | FL015671 | Heterocapsa triquetra | Species |
449_C | SA127154 | FL015671 | 449 | Strain |
449_C | SA127154 | FL015671 | Dinoflagellate | Broad taxon |
Collection:
Collection ID: | CO001583 |
Collection Summary: | Axenic phytoplankton were cultured in controlled laboratory settings and harvested under exponential growth using a gentle vacuum filtration onto 47 mm Durapore filters (pore size 0.2 µm). Samples were flash frozen in liquid N2 and stored at -80°C until extraction for metabolites. |
Sample Type: | Cultured plankton cells |
Storage Conditions: | Described in summary |
Treatment:
Treatment ID: | TR001603 |
Treatment Summary: | Media, light, and temperature were chosen for optimal growth of each phytoplankton species. In short, axenic phytoplankton were cultured in controlled laboratory settings and harvested under exponential growth using a gentle vacuum filtration onto 47 mm Durapore filters (pore size 0.2 µm). Samples were flash frozen in liquid N2 and stored at -80°C until extraction. In addition to samples, media blanks corresponding to each media type were harvested and served as a matrix blank to each corresponding phytoplankton sample. |
Sample Preparation:
Sampleprep ID: | SP001596 |
Sampleprep Summary: | Each sample was extracted using a modified Bligh-Dyer extraction. Briefly, filters were cut up and put into 15 mL teflon centrifuge tubes containing a mixture of 100 µm and 400 µm silica beads. Heavy isotope-labeled internal standards were added along with ~2 mL of cold aqueous solvent (50:50 methanol:water) and ~3 mL of cold organic solvent (dichloromethane). The samples were shaken on a FastPrep-24 Homogenizer for 30 seconds and chilled in a -20°C freezer repeatedly for three cycles of bead-beating and a total of 30 minutes of chilling. The organic and aqueous layers were separated by spinning samples in a centrifuge at 4,300 rpm for 2 minutes at 4°C. The aqueous layer was removed to a new glass centrifuge tube. The remaining organic fraction was rinsed three more times with additions of 1 to 2 mL of cold 50:50 methanol:water. All aqueous rinses were combined for each sample and ~2mL of cold dichloromethane was added to the combined aqueous layer. Tubes were shaken and centrifuged at 4,300 rpm for 2 minutes at 4°C. The aqueous layer was removed to a new glass vial and dried under N2 gas. The remaining organic layer in the bead beating tubes was transferred into the glass centrifuge tube and the bead beating tube was rinsed two more times with cold organic solvent. The combined organic rinses were centrifuged, transferred to a new glass vial, and dried under N2 gas. Dried aqueous fractions were re-dissolved in 380 µL of water. Dried organic fractions were re-dissolved in 380 µL of 1:1 water:acetonitrile. 20 µL of isotope-labeled injection standards in water were added to both fractions. Media blanks were extracted alongside samples as methodological blanks. |
Processing Storage Conditions: | On ice |
Extraction Method: | Bligh-Dyer |
Extract Storage: | -80℃ |
Combined analysis:
Analysis ID | AN002508 | AN002509 | AN002510 |
---|---|---|---|
Analysis type | MS | MS | MS |
Chromatography type | Reversed phase | HILIC | HILIC |
Chromatography system | Waters Acquity I-Class | Waters Acquity I-Class | Waters Acquity I-Class |
Column | Waters Acquity UPLC HSS Cyano (100 x 2.1mm,1.8um) | SeQuant ZIC-pHILIC (150 x 2.1mm,5um) | SeQuant ZIC-pHILIC (150 x 2.1mm,5um) |
MS Type | ESI | ESI | ESI |
MS instrument type | Orbitrap | Orbitrap | Orbitrap |
MS instrument name | Thermo Q Exactive HF hybrid Orbitrap | Thermo Q Exactive HF hybrid Orbitrap | Thermo Q Exactive HF hybrid Orbitrap |
Ion Mode | POSITIVE | POSITIVE | NEGATIVE |
Units | Adjusted and normalized peak areas | Adjusted and normalized peak areas | Adjusted and normalized peak areas |
Chromatography:
Chromatography ID: | CH001833 |
Chromatography Summary: | See attached summary. |
Methods Filename: | Ingalls_Lab_LC_Methods.txt |
Instrument Name: | Waters Acquity I-Class |
Column Name: | Waters Acquity UPLC HSS Cyano (100 x 2.1mm,1.8um) |
Column Temperature: | 35 |
Flow Rate: | 0.4 ml/min |
Solvent A: | 100% water; 0.1% formic acid |
Solvent B: | 100% acetonitrile; 0.1% formic acid |
Chromatography Type: | Reversed phase |
Chromatography ID: | CH001834 |
Chromatography Summary: | See attached summary |
Methods Filename: | Ingalls_Lab_LC_Methods.txt |
Instrument Name: | Waters Acquity I-Class |
Column Name: | SeQuant ZIC-pHILIC (150 x 2.1mm,5um) |
Column Temperature: | 30 |
Flow Rate: | 0.15 ml/min |
Solvent A: | 85% acetonitrile/15% water; 10 mM ammonium carbonate |
Solvent B: | 15% acetonitrile/85% water; 10 mM ammonium carbonate |
Chromatography Type: | HILIC |
MS:
MS ID: | MS002326 |
Analysis ID: | AN002508 |
Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | See attached protocol. |
Ion Mode: | POSITIVE |
Analysis Protocol File: | Ingalls_Lab_MS_Methods.txt |
MS ID: | MS002327 |
Analysis ID: | AN002509 |
Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | See attached protocol. |
Ion Mode: | POSITIVE |
Analysis Protocol File: | Ingalls_Lab_MS_Methods.txt |
MS ID: | MS002328 |
Analysis ID: | AN002510 |
Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | See attached protocol. |
Ion Mode: | NEGATIVE |
Analysis Protocol File: | Ingalls_Lab_MS_Methods.txt |